Methods and procedures for ligament repair

ABSTRACT

Methods and devices for the repair of a ruptured ligament using a scaffold device are provided. Aspects of the invention, may include a scaffold attached by a suture to an anchor. In aspects of the invention, the anchor may be secured to a bone near or at the repair site.

FIELD OF THE INVENTION

The invention relates generally to methods and devices for the repair ofa ruptured ligament using a scaffold device.

BACKGROUND OF THE INVENTION

Intra-articular tissues, such as the anterior cruciate ligament (ACL),do not heal after rupture. In addition, the meniscus and the articularcartilage in human joints also often fail to heal after an injury.Tissues found outside of joints heal by forming a fibrin clot, whichconnects the ruptured tissue ends and is subsequently remodeled to formscar, which heals the tissue. Inside a synovial joint, a fibrin cloteither fails to form or is quickly lysed after injury to the knee, thuspreventing joint arthrosis and stiffness after minor injury. Jointscontain synovial fluid which, as part of normal joint activity,naturally prevent clot formation in joints. This fibrinolytic processresults in premature loss of the fibrin clot scaffold and disruption ofthe healing process for tissues within the joint or withinintra-articular tissues.

The current treatment method for human anterior cruciate ligament repairafter rupture involves removing the ruptured fan-shaped ligament andreplacing it with a point-to-point tendon graft (ACL reconstruction).While this procedure can initially restore gross stability in mostpatients, longer follow-up demonstrates many post-operative patientshave abnormal structural laxity, suggesting the reconstruction may notwithstand the physiologic forces applied overtime (Dye, 325 Clin.Orthop. 130-139 (1996)). The loss of anterior cruciate ligament functionhas been found to result in early and progressive radiographic changesconsistent with joint deterioration (Hefti et al., 73A(3) J. Bone JointSurg. 373-383 (1991)), and over 70% of patients undergoing ACLreconstruction develop osteoarthritis at only 14 years after injury (vonPorat et al., Ann Rheum Dis. 63(3):269-73 (2004)). As anterior cruciateligament rupture is most commonly an injury of a young athletes in theirteens and twenties, early osteoarthritis in this group has difficultconsequences.

SUMMARY OF THE INVENTION

The invention relates in some aspects to methods and products thatfacilitate anterior cruciate ligament regeneration or healing. Thus, insome aspects the invention is a device for repairing a ruptured ligamenthaving a scaffold configured for repair of a ruptured ligament and ananchor. The scaffold is attached to the anchor with a suture. The suturehas at least one free end emerging from the scaffold. The suture and/oranchor may be bioabsorbable and/or synthetic, such as, for instance,polyglactin 910.

In some embodiments the scaffold is made of protein, such as, forexample, a synthetic, bioabsorbable, or a naturally occurring protein.In other embodiments the scaffold is a lyophilized material. Thescaffold may be expandable. In other embodiments the scaffold may be asponge, a gel, a solid, or a semi-solid. The scaffold may be pretreatedwith a repair material. Repair materials include but are not limited togels, liquids, and hydrogels. The repair material in some embodiments iscollagen.

A method of repairing a ruptured ligament is provided according to otheraspects of the invention. The method involves inserting a device forrepairing a ruptured ligament as described herein into a repair site ofthe ruptured ligament, attaching the anchor to a bone near the repairsite, and attaching the free end of the suture to an end of the rupturedligament.

A method of repairing a ruptured ligament that involves drilling a holenear a repair site of a ruptured ligament, attaching a suture to thebone through the hole, and attaching a scaffold to the suture to securethe scaffold between the bone and an end of the ruptured ligament isprovided in other aspects of the invention.

In some embodiments both ends of the suture are attached to the end ofthe ruptured ligament. In other embodiments the suture is attached to asecond bone site by a second anchor.

The scaffold in some embodiments is made from a protein. The protein maybe synthetic, bioabsorbable, or a naturally occurring protein. In someembodiments the scaffold can absorb plasma, blood, or other body fluids.

In other embodiments the scaffold is tubular, semi-tubular, cylindrical,or square. The scaffold is a sponge or a gel in some embodiments. Inother embodiments the scaffold is a semi-solid or, alternatively, asolid.

In yet other embodiments the scaffold is expandable. It may optionallyfill the repair site. In some embodiments the scaffold is bigger thanthe repair site and in other embodiments the scaffold partially fillsthe repair site. The scaffold may form around the ligament at the repairsite. The scaffold may be pretreated with a repair material, such as agel or a liquid. In some embodiments the repair material is a hydrogel.In other embodiments the repair material is collagen.

In some embodiments the ligament is ACL and the bone is a femur or atibia. In some embodiments the repair is supplemented by forming holesin the surrounding bone to cause bleeding into the repair site.

A method of repairing a ruptured ligament that involves drilling a holenear a repair site of a ruptured ligament and attaching an anchor to thebone through the hole is provided in some aspects of the invention. Themethod involves attaching an anchor to the bone through the hole wherethe anchor is attached to a scaffold and the scaffold is secured betweenthe bone and an end of the ruptured ligament.

In some embodiments, the ligament is ACL and the bone is a femur or atibia. In some embodiments, the anchor is bioabsorbable, metal, plastic,etc. In other embodiments, the anchor is a screw. In certainembodiments, the anchor is attached to the bone by a suture.

In some embodiments, the suture is a bioabsorbable, synthetic etc. Inother embodiments, the suture is polyglactin 910.

In some embodiments, the scaffold is synthetic, bioabsorbable, or anaturally occurring protein. In certain embodiments, the scaffold canabsorb plasma, blood, or other body fluids. In other embodiments, thescaffold is tubular, semi-tubular, cylindrical, or square. In certainembodiments, the scaffold is pretreated with a repair material. In someembodiments, the repair material is a gel or a liquid. In otherembodiments, the repair material is hydrogel. In some embodiments, therepair material is collagen.

In some embodiments, the repair is supplemented by forming holes in thesurrounding bone to cause bleeding into the repair site. In certainembodiments, the scaffold is expandable. It may optionally fill therepair site. In some embodiments the scaffold is bigger than the repairsite and in other embodiments the scaffold partially fills the repairsite. The scaffold may form around the ligament at the repair site. Thescaffold may be pretreated with a repair material, such as a gel or aliquid. In some embodiments the repair material is a hydrogel. In otherembodiments the repair material is collagen.

In some embodiments, the scaffold is a sponge. In certain embodiments,the scaffold is a gel. In other embodiments, the scaffold is asemi-solid. In some embodiments, the scaffold is a solid.

Each of the limitations of the invention can encompass variousembodiments of the invention. It is, therefore, anticipated that each ofthe limitations of the invention involving any one element orcombinations of elements can be included in each aspect of theinvention. This invention is not limited in its application to thedetails of construction and the arrangement of components set forth inthe following description or illustrated in the drawings. The inventionis capable of other embodiments and of being practiced or of beingcarried out in various ways. Also, the phraseology and terminology usedherein is for the purpose of description and should not be regarded aslimiting. The use of “including”, “comprising”, or “having”,“containing”, “involving”, and variations thereof herein, is meant toencompass the items listed thereafter and equivalents thereof as well asadditional items.

BRIEF DESCRIPTION OF THE DRAWINGS

The figures are illustrative only and are not required for enablement ofthe invention disclosed herein.

FIG. 1. A) Diagrammatic representation of a torn anterior cruciateligament. B) Diagrammatic representation of a scaffold device having ananchor and attached sutures. C) Diagrammatic representation of ascaffold device implanted into a repair site around a ruptured ACL.

FIG. 2. Diagrammatic representation of a method for inserting a scaffolddevice into bone. A) Diagrammatic representation of a suture anchorinserted into a bone. B) Diagrammatic representation of a drill hole ina bone and sutures attached to the opposite surface of the bone. C)Diagrammatic representation of a staple affixing a suture into a notch.D) Diagrammatic representation of an anchor with a central hole to allowbone marrow bleeding to flow into the attached scaffold. E) Diagrammaticrepresentation of an anchor with a scaffold sponge swaged directly ontoit.

FIG. 3. Diagrammatic representation of a method for distal fixation of ascaffold device to bone. A) Diagrammatic representation of a sutureattached through a drill hole in a bone. B) Diagrammatic representationof an anchor inserted into a bone.

FIG. 4. A) MRI image of ACL treated with sutures alone. B) MRI image ofACL treated with sutures+hydrogel. C) Diagrammatic representation of ACLwith suture only. D) Diagrammatic representation of ACL withsutures+hydrogel.

FIG. 5. A) MRI image of ACL treated with suture alone in the early, mid(5C) or late stage (E). B) MRI image of ACL treated with suture+hydrogelin the early, mid (SD) or late stage (F).

FIG. 6. A) MRI image of ACL scar treated with suture alone. B) MRI imageof ACL scar treated with suture+hydrogel.

FIG. 7. A) Photographic representation of ACL treated with suture alone.B) Photographic representation of ACL treated with suture+hydrogel.

FIG. 8. A) MRI image of intact ACL. B) ACL repaired with suture, anchorand sponge.

FIG. 9: Graph depicting biomechanical properties of Suture Anchor/SpongeRepair vs the current standard of care for ACL injuries (ACLReconstruction or ACLR) at 3 months in vivo.

DETAILED DESCRIPTION OF THE INVENTION

Aspects of the invention relate to devices and methods for repairing aruptured ligament. A device of the invention for the repair of aruptured ligament includes a scaffold which is configured for the repairof a ruptured ligament, an anchor and may include a suture. The scaffoldallows the subject's body to develop a network of capillaries, arteries,and veins. Well-vascularized connective tissues heal as a result ofmigration of fibroblasts into the scaffold. A device of the inventionprovides a connection between a ruptured ligament, or forms around atorn ligament, and promotes the repair of the ruptured or torn ligamentwhile maintaining the integrity and structure of the ligament.

The device of the invention provides a three-dimensional (3-D) scaffoldfor repairing a ruptured or torn ligament. The scaffold provides aconnection between the ruptured ends of the ligament and fibers, orforms around a torn ligament, after injury, and encourages the migrationof appropriate healing cells to form scar and new tissue in thescaffold. The scaffold is a bioengineered substitute for a fibrin clotand is implanted, for example, between the ruptured ends of the ligamentfascicles, or placed around a torn ligament. This substitute scaffold isdesigned to stimulate cell proliferation and extracellular matrixproduction in the gap between the ruptured ends of the ligament or thetear in the ligament, thus facilitating healing and regeneration.

Methods and devices of the invention may be used to treat eitherintra-articular or extra-articular injuries in a subject.Intra-articular injuries include, but are not limited to, meniscaltears, ligament tears and cartilage lesion. Extra-articular injuriesinclude, but are not limited to, the ligament, tendon or muscle. Thus,the methods of the invention may be used to treat injuries to theanterior cruciate ligament, the meniscus, labrum, for example glenoidlabrum and acetabular labrum, cartilage, and other tissues exposed tosynovial fluid after injury.

An injury may be a torn or ruptured ligament. A torn ligament is onewhere the ligament remains connected but has been damaged causing a tearin the ligament. The tear may be of any length or shape. A rupturedligament is one where the ligament has been completely severed providingtwo separate ends of the ligament. A ruptured ligament may provide twoligament ends of similar or different lengths. The rupture may be suchthat a ligament stump is formed at one end.

An example of a ruptured anterior cruciate ligament is depicted in FIG.1A. The anterior cruciate ligament (ACL) (2) is one of four strongligaments that connects the bones of the knee joint. The function of theACL is to provide stability to the knee and minimize stress across theknee joint. It restrains excessive forward movement of the lower legbone, the tibia (6), in relation to the thigh bone, the femur (4), andlimits the rotational movements of the knee. An anterior cruciateligament (2) is ruptured such that it no longer forms a connectionbetween the femur bone (4) and the tibia bone (6). The resulting ends ofthe ruptured ACL may be of any length. The ends may be of a similarlength, or one end may be longer in length than the other.

A scaffold of the device of the invention can be any shape that isuseful for implantation into a subject. The scaffold, for instance, canbe tubular, semi-tubular, cylindrical, including either a solid cylinderor a cylinder having hollow cavities, a tube, a flat sheet rolled into atube so as to define a hollow cavity, liquid, an amorphous shape whichconforms to that of the repair space, a “Chinese finger trap” design, atrough shape, or square.

Other shapes suitable for the scaffold of the device as known to thoseof ordinary skill in the art are also contemplated in the invention.

In aspects of the invention, a device for repairing a ruptured or tornligament includes a scaffold and an anchor, such that the scaffold isconfigured for repair. A scaffold that is configured for repair is onethat is capable of being inserted into an area requiring repair andpromotes regeneration of the ligament. A scaffold of the invention iscapable of insertion into a repair site and either forming a connectionbetween the ends of a ruptured ligament, or forming around a tornligament such that, in either case, the integrity and structure of theligament is maintained. Regeneration offers several advantages overreconstruction, previously used in ligament repair, includingmaintenance of the complex insertion sites and fan-shape of theligament, and preservation of remaining proprioceptive fibers within theligament substance.

Examples of devices and systems useful according to the invention aredepicted in FIGS. 1-3. An example of a device is depicted in FIGS. 1Band 1C. For example, a scaffold (14) is attached to a suture (12) and ananchor (8). The anchor (8) may, as shown in FIGS. 1B and 1C, be attachedto the suture (12) through an eyelet (10) of the anchor (8). The anchor(8) is attached (12) into a bone such as the femur (4) or a tibia (6).

A scaffold (14) may function either as an insoluble or biodegradableregulator of cell function or simply as a delivery vehicle of asupporting structure for cell migration or synthesis. Numerous matricesmade of either natural or synthetic components have been investigatedfor use in ligament repair and reconstruction. Natural matrices are madefrom processed or reconstituted tissue components (such as collagens andGAGs). Because natural matrices mimic the structures ordinarilyresponsible for the reciprocal interaction between cells and theirenvironment, they act as cell regulators with minimal modification,giving the cells the ability to remodel an implanted material, which isa prerequisite for regeneration.

Synthetic matrices are made predominantly of polymeric materials.Synthetic matrices offer the advantage of a range of carefully definedchemical compositions and structural arrangements. Some syntheticmatrices are not degradable. While the non-degradable matrices may aidin repair, non-degradable matrices are not replaced by remodeling andtherefore cannot be used to fully regenerate ligament. It is alsoundesirable to leave foreign materials permanently in a joint due to theproblems associated with the generation of wear particles, thusdegradable materials are preferred for work in regeneration. Degradablesynthetic scaffolds can be engineered to control the rate ofdegradation.

A scaffold is preferably made of a compressible, resilient materialwhich has some resistance to degradation by synovial fluid. Synovialfluid as part of normal joint activity, naturally prevents clotformation. This fibrinolytic process would result in the prematuredegradation of the scaffold and disrupt the healing process of theligament. The material may be either permanent or biodegradablematerial, such as polymers and copolymers. The scaffold can be composed,for example, of collagen fibers, collagen gel, foamed rubber, naturalmaterial, synthetic materials such as rubber, silicone and plastic,ground and compacted material, perforated material, or a compressiblesolid material.

A scaffold may be a solid material such that its shape is maintained, ora semi-solid material capable of altering its shape and or size. Ascaffold may be made of expandable material allowing it to contract orexpand as required. The material can be capable of absorbing plasma,blood, other body fluids, liquid, hydrogel, or other material thescaffold either comes into contact with or is added to the scaffold.

A scaffold material can be protein, lyophilized material, or any othersuitable material. A protein can be synthetic, bioabsorbable or anaturally occurring protein. A protein includes, but is not limited to,fibrin, hyaluronic acid, elastin, extracellular matrix proteins, orcollagen. A scaffold material may be plastic or self-assemblingpeptides. A scaffold material may incorporate therapeutic proteinsincluding, but not limited to, hormones, cytokines, growth factors,clotting factors, anti-protease proteins (e.g., alpha1-antitrypsin),angiogenic proteins (e.g., vascular endothelial growth factor,fibroblast growth factors), antiangiogenic proteins (e.g., endostatin,angiostatin), and other proteins that are present in the blood, bonemorphogenic proteins (BMPs), osteoinductive factor (IFO), fibronectin(FN), endothelial cell growth factor (ECGF), cementum attachmentextracts (CAE), ketanserin, human growth hormone (HGH), animal growthhormones, epidermal growth factor (EGF), interleukin-1 (IL-1), humanalpha thrombin, transforming growth factor (TGF-beta), insulin-likegrowth factor (IGF-1), platelet derived growth factors (PDGF),fibroblast growth factors (FGF, bFGF, etc.), and periodontal ligamentchemotactic factor (PDLLGF), for therapeutic purposes. A lyophilizedmaterial is one that is capable of swelling when liquid, gel or otherfluid is added or comes into contact with it.

Many biological materials are available for making the scaffold,including collagen compositions (either collagen fiber or collagen gel),compositions containing glycosaminoglycan (GAG), hyaluran compositions,and various synthetic compositions. Collagen-glycosaminoglycan (CG)copolymers have been used successfully in the regeneration of dermis andperipheral nerve. Porous natural polymers, fabricated as sponge-like andfibrous scaffolds, have been investigated as implants to facilitateregeneration of selected musculoskeletal tissues including ligaments. Ascaffold, such as a sponge scaffold, may also be made from tendon(xenograft, allograft, autograft) or ligament or skin or otherconnective tissue which could be in the native state or processed tofacilitate cell ingrowth or other biologic features.

In aspects of the invention, a scaffold is composed of a sponge orsponge-like material. A sponge scaffold may be absorbable ornonabsorbable. A sponge scaffold may be collagen, elastin, extracellularmatrix protein, plastic, or self-assembling peptides. A sponge scaffoldmay be hydrophillic. A sponge scaffold is capable of compression andexpansion as desired. For example, a sponge scaffold may be compressedprior to or during implantation into a repair site. A compressed spongescaffold allows for the sponge scaffold to expand within the repairsite. A sponge may be lyophilized and/or compressed when placed in therepair site and expanded once in place. The expansion of a spongescaffold may occur after contact with blood or other fluid in the repairsite or added to the repair site. A sponge scaffold may be porous. Asponge scaffold may be saturated or coated with a liquid, gel, orhydrogel repair material prior to implantation into a repair site.Coating or saturation of a sponge scaffold may ease implantation into arelatively undefined defect area as well as help to fill a particularlylarge defect area. A sponge scaffold may be composed of collagen. In apreferred embodiment, a sponge scaffold is treated with hydrogel.Examples of scaffolds and repair materials useful according to theinvention are found in U.S. Pat. No. 6,964,685 and US Patent ApplicationNos. 2004/0059416 and 2005/0261736, the entire contents of each areherein incorporated by reference.

An important subset of natural matrices are those made predominantlyfrom collagen, the main structural component in ligament. Collagen canbe of the soluble or the insoluble type. Preferably, the collagen issoluble, e.g., acidic or basic. For example, the collagen can be type I,II, III, IV, V, IX or X. Preferably the collagen is type I. Morepreferably the collagen is soluble type I collagen. Type I collagen isthe predominant component of the extracellular matrix for the humananterior cruciate ligament and provides an example of a choice for thebasis of a bioengineered scaffold. Collagen occurs predominantly in afibrous form, allowing design of materials with very differentmechanical properties by altering the volume fraction, fiberorientation, and degree of cross-linking of the collagen. The biologicproperties of cell infiltration rate and scaffold degradation may alsobe altered by varying the pore size, degree of cross-linking, and theuse of additional proteins, such as glycosaminoglycans, growth factors,and cytokines. In addition, collagen-based biomaterials can bemanufactured from a patient's own skin, thus minimizing the antigenicityof the implant (Ford et al., 105 Laryngoscope 944-948 (1995)).

A device of the invention may also include one or more anchors. Ananchor is a device capable of insertion into a bone such that it forms astable attachment to the bone. In some instances the anchor is capableof being removed from the bone if desired. An anchor may be conicalshaped having a sharpened tip at one end and a body having alongitudinal axis. The body of an anchor (8) may increase in diameteralong its longitudinal axis. The body of an anchor may include groovessuitable for screwing the anchor into position. For example, as depictedin FIG. 1C, the anchor (8) is screwed into the femur bone (4). An anchormay include an eyelet (10) at the base of the anchor body through whichone or more sutures may be passed. The eyelet (10) may be oval or roundand may be of any size suitable to allow one or more sutures to passthrough and be held within the eyelet (10).

An anchor may be attached to a bone by physical or mechanical methods asknown to those of ordinary skill in the art. An anchor includes, but isnot limited to, a screw, a barb, a helical anchor, a staple, a clip, asnap, a rivet, or a crimp-type anchor. The body of an anchor may bevaried in length. Examples of anchors, include but are not limited to,IN-FAST™ Bone Screw System (Influence, Inc., San Francisco, Calif.),IN-TAC™ Bone Anchor System (Influence, Inc., San Francisco, Calif.),Model 3000 AXYALOOP™ Titanium Bone Anchor (Axya Medical Inc., Beverly,Mass.), OPUS MAGNUM® Anchor with Inserter (Opus Medical, Inc., San JuanCapistrano, Calif.), ANCHRON™, HEXALON™, TRINION™ (all available fromInion Inc., Oklahoma City, Okla.) and TwinFix AB absorbable sutureanchor (Smith & Nephew, Inc., Andover, Mass.). Anchors are availablecommercially from manufacturers such as Influence, Inc., San Francisco,Calif., Axya Medical Inc., Beverly, Mass., Opus Medical, Inc., San JuanCapistrano, Calif., Inion Inc., Oklahoma City, Okla., and Smith &Nephew, Inc., Andover, Mass.

An anchor may be attached directly to a scaffold where the anchor isswaged directly onto the scaffold. FIG. 2E depicts such an example. Theanchor (8) is attached directly to the scaffold (14) by its base end andthe anchor (8) is attached to the femur (4) by its sharpened end.

An anchor may be attached indirectly to a scaffold using a suture tosecure it in position. FIG. 2A depicts such an example. A suture (12) ispassed through the eyelet (10) of the anchor (8) and held within theeyelet (10) to attach the scaffold (14). The first end (16) and thesecond end (18) of the suture are free and emerge from the scaffold(14). The anchor (8) is attached to the femur (4) by its sharpened end.

An anchor may be composed of a non-degradable material, such as metal,for example titanium 316 LVM stainless steel, CoCrMo alloy, or Nitinolalloy, or plastic. An anchor is preferably bioabsorbable such that thesubject is capable of breaking down the anchor and absorbing it.Examples of bioabsorbable material include, but are not limited to,MONOCRYL (poliglecaprone 25), PDS II (polydioxanone), surgical gutsuture (SGS), gut, coated VICRYL (polyglactin 910, polyglactin 910braided), human autograft tendon material, collagen fiber, POLYSORB,poly-L-lactic acid (PLLA), polylactic acid (PLA), polysulfone,polylactides (Pla), racemic form of polylactide (D,L-Pla),poly(L-lactide-co-D,L-lactide), 70/30 poly(L-lactide-co-D,L-lactide),polyglycolides (PGa), polyglycolic acid (PGA), polycaprolactone (PCL),polydioxanone (PDS), polyhydroxyacids, and resorbable plate material(see e.g. Orthopedics, October 2002, Vol. 25, No. 10/Supp.). The anchormay be bioabsorbed over a period of time which includes, but is notlimited to, days, weeks, months or years.

An anchor may have a central hole (24) through which fluids, such asblood, may pass. The hole (24) may allow such fluids to flow onto theattached scaffold. FIG. 2D depicts such an example. The anchor (8) isattached to the femur (4) and includes a central hole (24) through whichblood can pass. Blood is able to pass through the central hole (24) inthe anchor (8) and onto the scaffold (14) which absorbs the blood.

In aspects of the invention, an anchor (8) may be attached to a scaffold(14) using a suture (12). FIG. 1B illustrates an example of an anchorattached to a scaffold using a suture. A suture (12) is passed throughthe eyelet (10) of an anchor (8) such that the anchor (8) is attached tothe scaffold (14) by the suture (12). The suture (12) has at least onefree end. In some embodiments, a suture has two free ends, a first end(16) and a second end (18).

A suture (12) is preferably bioabsorbable, such that the subject iscapable of breaking down the suture and absorbing it, and synthetic suchthat the suture may not be from a natural source. A suture (12) may bepermanent such that the subject is not capable of breaking down thesuture and the suture remains in the subject. A suture (12) may be rigidor stiff, or may be stretchy or flexible. A suture (12) may be round inshape and may have a flat cross section. Examples of sutures include,but are not limited to, VICRYL™ polyglactin 910, PANACRYL™ absorbablesuture, ETHIBOND® EXCEL polyester suture, PDS® polydioxanone suture andPROLENE® polypropylene suture. Sutures are available commercially frommanufacturers such as MITEK PRODUCTS division of ETHICON, INC. ofWestwood, Mass.

A suture (12) may be attached to one or both ends of a ruptured ligamentby its first end (16) and/or its second end (18). FIG. 1C illustrates anexample of a device of the invention inserted into a repair site of aruptured ligament. A suture (12) is passed through the eyelet (10) ofthe anchor and the first end (16) and second end (18) are tied to theends of the distal ACL (2). The anchor (8) is attached to the femur (4)by its sharpened end. The scaffold (14) attached to the anchor (8) bythe suture (12) is held in position in the repair site (26). The anchor(8) may be attached to either the tibia bone (6) or the femur bone (4)to secure the scaffold (14) in position.

A staple (22) is a type of anchor having two arms that are capable ofinsertion into a bone. In some instances, the arms of the staple fold inon themselves when attached to a bone or in some instances when attachedto other tissue. A staple may be composed of metal, for example titaniumor stainless steel, plastic, or any biodegradable material. A stapleincludes but is not limited to linear staples, circular staples, curvedstaples or straight staples. Staples are available commercially frommanufacturers such as Johnson & Johnson Health Care Systems, Inc.Piscataway, N.J., and Ethicon, Inc., Somerville, N.J. A staple may beattached using any staple device known to those of ordinary skill in theart, for example, a hammer and staple setter (staple holder).

In some embodiments, a staple may be used to hold the suture securely inposition. A suture may be attached to a bone using a staple as depictedin FIG. 2C. A suture (12) is held in place in the femur (4) with astaple (22) such that the first end (16) and the second end (18) of thesuture (12) are free.

Aspects of the invention relate to methods of repairing a ruptured ortorn ligament. In some embodiments, a device of the invention isinserted into a repair site of the ruptured or torn ligament. In certainembodiments, a hole is drilled into a bone at or near a repair site of aruptured or torn ligament and a suture is attached through the hole tothe bone.

A repair site (26) is the area around a ruptured or torn ligament (2)into which a device of the invention may be inserted. A device of theinvention may be placed into a repair site (26) area during surgeryusing techniques known to those of ordinary skill in the art. A scaffold(14) of the invention can either fill the repair site (26) or partiallyfill the repair site (26). A scaffold (14) can partially fill the repairsite (26) when inserted and expand to fill the repair site (26) in thepresence of blood, plasma or other fluids either present within therepair site (26) or added into the repair site (26).

A scaffold (14) may form around a ruptured or torn ligament (2) at therepair site (26). For example, a scaffold (14) may be formed into a tubeshape and wrapped around a ligament, a scaffold (14) may be positionedbehind the ligament such that the ligament is held within the scaffold(14), or a scaffold (14) may be a “Chinese finger trap” design where oneend is placed over a stump of a ruptured ligament and the second endplaced over the other end of the ruptured ligament.

Aspects of the invention provide methods of repairing a rupturedligament (2) involving drilling a hole (20) at or near a repair site(26) of a ruptured ligament (2). A bone at or near a repair site is onethat is within close proximity to the repair site and can be utilizedusing the methods and devices of the invention. For example, a bone ator near a repair site of a torn anterior cruciate ligament is a femur(4) bone and/or a tibia (6) bone. A hole can be drilled into a boneusing a device such as a Kirschner wire (for example a small Kirschnerwire) and drill, or microfracture pics or awls. One or more holes may bedrilled into a bone surrounding a repair site to promote bleeding intothe repair site. The repair can be supplemented by drilling holes intothe surrounding bone to cause bleeding. Encouraging bleeding into therepair site may promote the formation of blood clots and enhance thehealing process of the injury.

A hole (20) may be drilled into a bone on the opposite side to therepair site (26). A suture (12) may be passed through the hole (20) inthe bone and attached to the bone. A scaffold (14) is attached to thesuture (12) to secure the scaffold (14) between the bone and an end of aruptured ligament (2). A ruptured ligament (2) provides two ends of theligament that were previously connected. A scaffold (14) may be attachedto one or both ends (16, 18) of a ruptured ligament (2) by one or moresutures (12). A suture (12) may be attached to a second bone site at ornear the repair site. The suture may be attached to the second boneusing a second anchor (8).

An example of such a method is depicted in FIG. 2B. A hole is drilled(20) into the opposite side of the femur bone (4). The suture (12) isattached to the opposite side of the femur bone (4) using the first end(16) and the second end (18) through the hole (20).

Another example is depicted in FIG. 3A. A hole (20) is drilled into thetibia (6) near the end of the ruptured ligament (2) and a suture isattached to the tibia (6) through the hole (20).

A scaffold of the device can be pretreated with a repair material priorto implantation into a subject. The scaffold may be soaked in a repairmaterial prior to or during implantation into a repair site. The repairmaterial may be injected directly into the scaffold prior to or duringimplantation. The repair material may be injected within a tubularscaffold at the time of repair. Repair material includes, but is notlimited to, a gel, for example a hydrogel, a liquid, or collagen. Aliquid includes any material capable of forming an aqueous material, asuspension or a solution. A repair material may include additionalmaterials, such as growth factors, antibiotics, insoluble or solublecollagen (in fibrous, gel, sponge or bead form), a cross-linking agent,thrombin, stem-cells, a genetically altered fibroblast, platelets,water, plasma, extracellular proteins and a cell media supplement. Theadditional repair materials may be added to affect cell proliferation,extracellular matrix production, consistency, inhibition of disease orinfection, tonicity, cell nutrients until nutritional pathways areformed, and pH of the repair material. All or a portion of theseadditional materials may be mixed with the repair material before orduring implantation, or alternatively, the additional materials may beimplanted proximate to the defect area after the repair material is inplace.

In certain embodiments, a repair material may include collagen andplatelets. In some embodiments, platelets are derived from the subjectto be treated. In other embodiments, platelets are derived from a donorthat is allogeneic to the subject. In certain embodiments, platelets maybe obtained as platelet rich plasma (PRP). In a non-limiting example,platelets may be isolated from a subject's blood using techniques knownto those of ordinary skill in the art. As an example, a blood sample maybe centrifuged at 700 rpm for 20 minutes and the platelet-rich plasmaupper layer removed. Platelet density may be determined using a cellcount as known to those of ordinary skill in the art. The platelet richplasma may be mixed with collagen and used as a scaffold. The plateletrich plasma may be mixed with any one or more of the scaffold materialsof the invention.

An example of a gel is a hydrogel. A hydrogel is a substance that isformed when an organic polymer (natural or synthetic) is crosslinked viacovalent, ionic, or hydrogen bonds to create a three-dimensionalopen-lattice structure which entraps water molecules to form a gel. Apolymer may be crosslinked to form a hydrogel either before or afterimplantation into a subject. For instance, a hydrogel may be formed insitu, for example, at a repair site. In certain embodiments, a polymerforms a hydrogel within the repair site upon contact with a crosslinkingagent. Naturally occurring and synthetic hydrogel forming polymers,polymer mixtures and copolymers may be utilized as hydrogel precursors.See for example, U.S. Pat. No. 5,709,854. In certain embodiments, ahydrogel is a gel and begins setting immediately upon mixture and takesapproximately 5 minutes to sufficiently set before closure of the defectand surgery area. Setting time may vary depending on the mixture of gelused and environmental factors.

For instance, certain polymers that can form ionic hydrogels which aremalleable may be used to form the hydrogel. For example, a hydrogel canbe produced by cross-linking the anionic salt of alginic acid, acarbohydrate polymer isolated from seaweed, with calcium cations, whosestrength increases with either increasing concentrations of calcium ionsor alginate. Modified alginate derivatives, for example, which have animproved ability to form hydrogels or which are derivatized withhydrophobic, water-labile chains, e.g., oligomers of ε-caprolactone, maybe synthesized. Additionally, polysaccharides which gel by exposure tomonovalent cations, including bacterial polysaccharides, such as gellangum, and plant polysaccharides, such as carrageenans, may be crosslinkedto form a hydrogel. Additional examples of materials which can be usedto form a hydrogel include polyphosphazines and polyacrylates, which arecrosslinked ionically, or block copolymers such as PLURONICS™(polyoxyalkylene ether) or TETRONICS™ (nonionic polymerized alkyleneoxide), polyethylene oxide-polypropylene glycol block copolymers whichare crosslinked by temperature or pH, respectively. Other materialsinclude proteins such as fibrin, polymers such as polyvinylpyrrolidone,hyaluronic acid and collagen. Polymers such as polysaccharides that arevery viscous liquids or are thixotropic, and form a gel over time by theslow evolution of structure, are also useful.

Another example of a gel is hyaluronic acid. Hyaluronic acid, whichforms an injectable gel with a consistency like a hair gel, may beutilized. Modified hyaluronic acid derivatives are particularly useful.Hyaluronic acid is a linear polysaccharide. Many of its biologicaleffects are a consequence of its ability to bind water, in that up to500 ml of water may associate with 1 gram of hyaluronic acid.Esterification of hyaluronic acid with uncharged organic moietiesreduces the aqueous solubility. Complete esterification with organicalcohols such as benzyl renders the hyaluronic acid derivativesvirtually insoluble in water, these compounds then being soluble only incertain aprotic solvents. When films of hyaluronic acid are made, thefilms essentially are gels which hydrate and expand in the presence ofwater.

A gel may be provided in pharmaceutical acceptable carriers known tothose skilled in the art, such as saline or phosphate buffered saline.Such carriers may routinely contain pharmaceutically acceptableconcentrations of salt, buffering agents, preservatives, compatiblecarriers, supplementary immune potentiating agents such as adjuvants andcytokines and optionally other therapeutic agents.

As used herein, the term “pharmaceutically acceptable” means a non-toxicmaterial that does not interfere with the effectiveness of thebiological activity of the scaffold material or repair material. Theterm “physiologically acceptable” refers to a non-toxic material that iscompatible with a biological system such as a cell, cell culture,tissue, or organism. The characteristics of the carrier will depend onthe route of administration. Physiologically and pharmaceuticallyacceptable carriers include diluents, fillers, salts, buffers,stabilizers, solubilizers, and other materials which are well known inthe art. The term “carrier” denotes an organic or inorganic ingredient,natural or synthetic, with which the scaffold material is combined tofacilitate the application. The components of the pharmaceuticalcompositions also are capable of being co-mingled with the device of thepresent invention, and with each other, in a manner such that there isno interaction which would substantially impair the desiredpharmaceutical efficacy.

The devices of the invention may be used in surgical procedures. Thefollowing is an example of a surgical procedure which may be performedusing the devices and methods of the invention. The affected extremityis prepared and draped in the standard sterile fashion. A tourniquet maybe used if indicated. Standard arthroscopy equipment may be used. Afterdiagnostic arthroscopy is performed, and the intra-articular lesionidentified and defined, the tissue ends are pretreated, eithermechanically or chemically, and the scaffold introduced into the tissuedefect. The scaffold is then bonded to the surrounding tissue using themethods described herein. This can be done by the addition of a chemicalagent or a physical agent such ultraviolet light, a laser, or heat. Thescaffold may be reinforced by placement of sutures or clips. Thearthroscopic portals can be closed and a sterile dressing placed. Thepost-operative rehabilitation is dependent on the joint affected, thetype and size of lesion treated, and the tissue involved.

The device of the invention may be used with arthroscopic equipment. Thedevice of the invention may be used by insertion through an openincision. The scaffold is compressible to allow introduction througharthroscopic portals, incisions and equipment. The scaffold can also bepre-treated in antibiotic solution prior to implantation.

A subject includes, but is not limited to, any mammal, such as human,non-human primate, mouse, rat, dog, cat, horse or cow. In certainembodiments, a subject is a human.

The invention also includes in some aspects kits for repair of rupturedor torn ligaments. A kit may include a scaffold of the invention havingat least one anchor attached to the scaffold and instructions for use.The scaffold may further include one or more sutures that attach ananchor to the scaffold. A kit may further include a container thatcontains a repair material as described herein.

EXAMPLES Example 1

Bilateral ACL transections were performed in six animals and repairedwith a four stranded, absorbable suture repair using a variation of theMarshall technique. For each animal, one of the repairs was augmentedwith placement of a collagen-platelet rich hydrogel at the ACLtransection site, while the contralateral knee had suture repair alone.No post-operative immobilization was used. The animals survived for fourweeks and then underwent in vivo magnetic resonance imaging followed byeuthanasia and immediate biomechanical testing. Six control knees withintact ACLs from three additional 30 kg pigs were also testedbiomechanically as an intact ACL control group.

The supplementation of suture repair with a collagen-platelet richhydrogel resulted in formation of a large scar mass in the region of theACL which was perfused by the injection of IV gadolinium, suggesting theformation of a vascularized repair tissue in the ACL defect.

Despite suture resorption during the in vivo time course, load at yield,stiffness and displacement at yield all improved when collagen-plateletrich hydrogel was used to augment the suture repairs. The use of suturerepair alone, or suture repair augmented with a collagen-platelet poorhydrogel did not show improvement in any of these parameters.

Biomechanical healing of the porcine ACL after complete transection andsuture repair can be enhanced at an early time point with use of acollagen-platelet rich hydrogel placed in the wound site at the time ofprimary repair.

Developing a technique for primary repair of the ACL may change thefocus of treatment of this injury from resection and reconstructiontoward repair and regeneration.

Example 2 Experimental Design

Seven 30 kg Yorkshire pigs underwent bilateral ACL transection andsuture repair.

Five of the animals were treated on one side with suture repair on oneside and suture repair augmented with collagen-platelet rich plasmacontaining an average of 954K+/−93K platelets/mm³ on the contralateralside (n=5). An additional two animals had suture repair on one side andsuture repair augmented with collagen-platelet poor plasma (n=2) with aplatelet counts less than 20 K/mm³ on the contralateral side (n=5).Sides were randomized to suture alone and augmented repair. All outcomeswere measured after four weeks in vivo. Just prior to euthanasia, theanimals had in vivo MRI of both knees with gadolinium contrast to assessperfusion of the ACL wound site. Immediately after euthanasia, the kneeswere harvested and ex vivo MRI performed, followed immediately bybiomechanical testing of the ACL complex as previously described(Murray, M. M.; Spindler, K. P.; Devin, C.; Snyder, R. B.; Muller, J.;Ballard, P.; Nanney, L. B.; and Zurakowski, D.: Healing of anintra-articular tissue defect using a stabilized provisional scaffold.Journal of Bone & Joint Surgery—American Volume, submitted forpublication, 2005). Intact ACLs (n=6) were used as a control group forthe biomechanical studies.

Manufacture of Acid Soluble Collagen Used in the Hydrogels:

Rat tails were obtained from control breeder rats undergoing euthanasia.The rat tail tendons were sterilely harvested, minced, and solubilizedin an acidified pepsin solution to obtain the acid soluble collagen.Collagen content within the slurry was adjusted to approximately 10mg/ml using a hydroxyproline assay and additional 0.01N HCl to adjustthe content as needed. Before combining with the platelet component ofthe hydrogel, the collagen slurry was mixed with 30% 0.1M HEPES, 20% 10×Ham's F10, 30% Penicillin/streptomycin/amphoptericin B and 30% sterilewater. The collagen slurry was then neutralized to a pH of 7.4 using7.5% sodium bicarbonate.

PRP Preparation:

Whole blood was drawn from the jugular vein of each pig into tubescontaining sodium citrate immediately prior to surgery. The blood wascentrifuged to isolate the platelet-rich plasma (PRP) fraction at 100 gfor 14 minutes. This resulted in an approximately 2× enrichment of theplatelet concentration of the blood from a range of 495 to 567 K/mm³ to780 to 2300 K/mm³. To make the platelet poor plasma (PPP), theanticoagulated blood was centrifuged at 200 g for twenty minutes,resulting in platelet counts of 18K/mm³ and 14K/mm³ in the two samples.The PRP or PPP was added to the collagen slurry to keep theplasma-collagen ratio at 1:1. The mixture was kept on ice until use.

Surgical Procedure:

IACUC approvals were obtained for this study prior to any surgicalprocedures. The pigs were pre-medicated with telazol 4.4-6.6 mg/kgintra-muscular (IM), xylazine 1.1-2.2 mg/kg IM, and atropine 0.04 mg/kg.They were intubated and placed on isoflurane 1-3% for anesthesiamaintenance. After anesthesia had been obtained, the pigs were weighedand range of motion of each knee measured using a goniometer. The pigswere then placed in the supine position on the operating room table.Both hind limbs were shaved, prepared with chlorhexidine followed bybetadyne paint and sterilely draped. To expose the ACL, afour-centimeter incision was made over the medial border of the patellartendon. The incision was carried down sharply through the synovium. Allsharp dissection after the skin incision was done using electrocauteryfor hemostasis. The fat pad was released from its proximal attachmentand partially resected to expose the intermeniscal ligament. Theintermeniscal ligament was released to expose the tibial insertion ofthe ACL. A Lachman maneuver was performed prior to releasing the ACL toverify knee stability. Two #1 Vicryl sutures were secured in the distalACL stump using a modified Kessler stitch and the ends clamped. The ACLwas transected completely using a No. 12 blade. Complete transection wasverified visually and with a repeat Lachman maneuver. An absorbablesuture anchor (TwinFix AB 5.0 Suture Anchor with DuraBraid Suture(USP#2); Smith & Nephew, Inc, Andover Mass.) was placed at the back ofthe femoral notch. The knee was irrigated with 500 cc of sterile normalsaline to remove all synovial fluid. Hemostasis was carefully achievedusing pressure and a solution of 1:10,000 of epinephrine as needed. Oncehemostasis had been achieved, a strip of Gelfoam was presoaked in onebatch of the collagen-PRP mixture, and threaded onto sutures and up intothe region of the proximal ACL stump in the notch. The sutures were tiedwith the knees in resting flexion (approximately 70 degrees) and asecond batch of the collagen-platelet mixture was placed on top of theGelfoam in the experimental knees. The knee was closed after the gelreached a soft set (approximately 10 minutes). The knee was left inresting extension while the suture repair alone was performed on thecontralateral knee (approximately 1 hour). The procedure was identicalin the suture repair alone knees, with the exception of the placement ofthe Gelfoam sponge and collagen-PRP hydrogel. In the collagen-PPP group,the procedure was identical to the PRP group, with the substitution ofthe platelet poor plasma for the platelet rich plasma in thecollagen-platelet hydrogel. The incisions were closed in layers.

The animals were not restrained post-operatively, and were allowed adlib activity. Once the animals recovered from anesthesia, they werepermitted to resume normal cage activity and nutrition ad lib. Banamine1.1 mg/kg IM once and a Fentanyl patch 1-4 ug/kg transdermal wereprovided for post-operative analgesia. All animals were weight bearingon their hind limbs by 24 hours after surgery. After four weeks in vivo,the animals were again anesthetized and underwent in vivo MR imagingusing the protocol detailed below.

After the magnetic resonance images had been obtained, the animals wereeuthanized using Fatal Plus at 1 cc/10 lbs. There were no animals whichhad any surgical complications or difficulty walking normally, redness,warmth and swelling of the knee, fever or other signs of infection thatwould have necessitated early euthanasia. The knees were retrieved andtaken for immediate ex vivo MR imaging and same-day biomechanicaltesting. The knees were kept at 4° C. until biomechanical testing andkept moist using a saline spray and moist wraps.

Magnetic Resonance Imaging

In vivo magnetic resonance imaging was performed at 1.5 Tesla (GEMedical Systems, Milwaukee, Wis.) with a dedicated surface coil at thespecified time points. Scanning was performed with the knees held in 30degrees of flexion. Conventional MR images included T1 and 3D FSE protondensity sequences. Multisection-multiechoT2 sequence for mapping T2relaxation time were obtained with the following imaging parameters:TRITE, 4000/14-98 in 14-msec increments for a total of 7 echo imageswith a 3-mm slice thickness. Perfusion was evaluated by using spoiledgradient echo sequence (TR/TE=200/2 ms, flip angle=60, 3 mm slicethickness, and 0.625 mm in plane resolution) with an intravenouscontrast agent (Magnevist; Berlex, Wayne, N.J.) injected 10 s after thestart of scan. Five images were obtained per slice, 78 s apart. Postcontrast T1-weighted images were obtained (TR/TE=500/9 ms) in thecoronal and sagittal planes with a 3-mm slice thickness.

Ex vivo magnetic resonance imaging was conducted on a 4.7 Teslamicroimaging system (Biospec™, Bruker BioSpin MRI, Inc., Karlsruhe,Germany). The system consisted of three-axis self-shielded magneticfield gradients, with 30 G/cm maximum gradient amplitude in all threechannels. The six-week treated and intact knee specimens were placed inthe radiofrequency coil (I.D. 72 mm) with the knee flexed 30 degrees asverified by goniometer. After the T2 localizers imaging on threeorthogonal axes, spin-echo proton density weighted images (PD-WIs) weresubsequently acquired in the sagittal and axial planes at the ACL site.The pulse sequence used was a conventional spin-echo sequence withfollowing parameters: repetition time (TR)=2 sec, echo time (TE)=10msec, band width=100 kHz, field of view=50 mm, matrix size=256×256,slice thickness=1 mm (gapless), number of slices=15, affording a totalscan time of approximately 8.5 minutes.

Biomechanical Testing

The bone-ligament-bone ACL complex from both knees for each pig wastested in uniaxial tension. After euthanasia, the hind limb of each pigwas amputated through the midshaft of the femur and tibia and the skinand overlying muscle removed. The joint capsule, MCL, lateral collateralligament and the posterior cruciate ligament were kept intact during theembedding process and preserved until just prior to mechanical testingto facilitate correct spacing and alignment of the femur and tibiaduring mounting of the knee in the mechanical test apparatus. The ACLwas maintained in a hydrated state throughout preparation and mechanicaltesting by wrapping the dissected knee in gauze and irrigatingrepeatedly with 0.9% saline. Temperature was maintained at roomtemperature to allow comparison with other published studies. The femurand tibia were cut to four inches in length and the ends embedded incylindrical molds using polymethylmethacrylate (PMMA) resin. One or twotransversely oriented, self-tapping drywall screws were placedunicortically in the proximal femur and distal tibia prior to embeddingto prevent inadvertent pullout of the femur and/or tibia from the PMMAduring. mechanical testing. A specially designed jig was used toposition the femur and tibia in the cylindrical molds with the kneeflexed to 30° to align the femoral and tibial attachments of the ACLcoaxial with the line of action of the load actuator. A goniometer wasused to measure the overall alignment and orientation of the femur,tibia and ACL complex before embedding. After approximately 30 minutescure time, the specimens are positioned in the grips and the remainingsoft tissue attachments sectioned so that only the ACL was capable ofresisting distraction across the knee joint during mechanical tensiletesting.

All mechanical testing was conducted using an Interlaken Series 3300Load frame (Eden Prairie, Minn.) controlled by an MTS TestStar IImDigital Controller (Eden Prairie, Minn.). All test parameters werepreprogrammed and all phases of testing were executed automatically soas to maintain consistency during testing of all the specimens. Once thespecimen was locked in the grips, the force and displacement transducerswere zeroed. Close-range digital images of the bone-ligament-bone ACLcomplex were acquired at 3 Hz using a high resolution digital camerawith a macro lens (PixeLINK PLA662 Megapixel Firewire camera, PixeLINK,Ottawa ON, Canada) so that the portion of the ACL that failed (i.e.midsubstance at repair site, femoral or tibial attachment sites) couldbe observed directly. Before conducting the tensile test, thebone-ligament-bone ACL complex was preconditioned with ten cycles ofloading and unloading at a strain amplitude of approximately 3%, at arate of 5 mm/min (Sakai, T.; Yasuda, K.; Tohyama, H.; Azuma, H.; Nagumo,A.; Majima, T.; and Frank, C. B.: Effects of combined administration oftransforming growth factor-beta1 and epidermal growth factor onproperties of the in situ frozen anterior cruciate ligament in rabbits.Journal of Orthopaedic Research, 20(6): 1345-51, 2002) to eliminate any“slack” in the test setup and to minimize viscoelastic effects (creepand stress-relaxation). Immediately after preconditioning, each specimenwas tested to failure in uniaxial tension at 20 mm/min (Sakai, T.;Yasuda, K.; Tohyama, H.; Azuma, H.; Nagumo, A.; Majima, T.; and Frank,C. B.: Effects of combined administration of transforming growthfactor-beta1 and epidermal growth factor on properties of the in situfrozen anterior cruciate ligament in rabbits. Journal of OrthopaedicResearch, 20(6): 1345-51, 2002; Katsuragi, R.; Yasuda, K.; Tsujino, J.;Keira, M.; and Kaneda, K.: The effect of nonphysiologically high initialtension on the mechanical properties of in situ frozen anterior cruciateligament in a canine model. American Journal of Sports Medicine, 28(1):47-56, 2000). The applied actuator displacement and resultant force datawere acquired at 10 Hz. After mechanical testing was completed, theproximal and distal portions of the ruptured ACL were explanted from thebone and submitted for further gross and microscopic analysis.

The tangent modulus (maximum slope of force-displacement curve), maximumload at failure and total work to failure (area under force-displacementcurve) were determined from the force-displacement curve measured foreach bone-ligament-bone ACL complex tested. Data was analyzed usingMATLAB (The Math Works, Natick, Mass.). The yield load represents thepoint along the normalized force-displacement curve where the mechanicalbehavior of the ACL complex departed from “linear” behavior and for thepurposes of this analysis was defined as the point where the tangentmodulus declines by at least 2% from its maximum value. The ultimateload was deduced from the maximal normalized load sustained by the ACLcomplex prior to failure. The work to failure was derived by integratingthe total area under the force-displacement curve.

Magnetic Resonance Imaging

In vivo MRI demonstrated a large mass of scar tissue in the area of theACL transections treated with suture and collagen-platelet hydrogel,with a smaller mass seen in the region of the ACL when sutures alonewere used (FIG. 4). In the knees treated with suture and thecollagen-platelet hydrogel, the tissue in the region of the ACL appearedto be developing linear densities consistent with collagen fascicleswithin the mass, coursing from femur to tibia. The fat pad anterior tothe scar mass in the knees treated with suture and collagen-platelethydrogel also enhanced strongly immediately on perfusion of the kneewith the IV gadolinium contrast, whereas the knee treated with suturerepair alone had less visible enhancement (FIG. 5). The difference inscar size and perfusion between the knees treated with suture alone andthe knees treated with suture and collagen-platelet hydrogel was seen onthe post-gadolinium coronal images as well (FIG. 6).

Biomechanics

Failure Mode: In the ligaments treated with absorbable suture repairalone or suture repair augmented with platelet poor hydrogel, the modeof failure was intra-substance in 6 out of 6 ligaments, while in thosetreated with suture repair+PRP-hydrogel, the repaired ligaments failedat the bone-ligament junction in 2 out of 5 cases (FIG. 7; Table 1). Inthe intact ligaments, failure was at the bone-ligament junction in 6 outof 6 cases. The absorbable suture material used in the suture repairsappeared to be completely resorbed at the four week time point.

Load at Yield: After four weeks in vivo, the suture repairs augmentedwith collagen-PRP hydrogel had a load at yield almost three times ashigh as the repairs performed with suture repair alone (Table 1). Theyield load of the collagen-PRP group reached 65% of the yield load ofthe intact ACLs (179+/−37N) during these four weeks in vivo. Thestrength of the suture repair+PRP-hydrogel was significantly greaterthan that of the suture repairs alone, but both groups were stillsignificantly lower than the intact ACL at four weeks (ANOVA, p<0.0001for group, Bonferroni-Dunn correction post-hoc testing p<0.008 for allcomparisons).

Maximum Load After four weeks in vivo, the suture repairs augmented withcollagen-PRP hydrogel held a maximum load that was twice as high as therepairs performed with suture repair alone (Table 1). The maximum loadof the collagen-PRP group reached 57% of the maximum load of the intactACLs (179+/−37N) during these four weeks in vivo. The strength of thesuture repair+PRP-hydrogel was significantly greater than that of thesuture repairs alone, but both groups were still significantly lowerthan the intact ACL at four weeks (ANOVA, p<0.0001 for group,Bonferroni-Dunn correction post-hoc testing p<0.008 for allcomparisons).

Displacement at Yield: While the mean value of displacement at yield waslower in the repairs augmented with collagen-PRP hydrogel than in thesuture repairs alone (Table 1), differences between the four groups werenot found to be significant (ANOVA, p>0.07 for group, p>0.008 for allcomparisons).

Stiffness: After four weeks in vivo, the stiffness of the suture repairsaugmented with collagen-PRP hydrogel was twice as high as the repairsperformed with suture repair alone (Table 1). The stiffness of thecollagen-PRP group reached 50% of the stiffness of the intact ACLsduring these four weeks in vivo. The stiffness of the suturerepair+PRP-hydrogel was significantly greater than that of the suturerepairs alone, but both groups were still significantly lower than theintact ACL at four weeks (ANOVA, p<0.0001 for group, Bonferroni-Dunncorrection post-hoc testing p<0.008 for all comparisons).

Energy to Failure: The energy to failure in the suturerepair+PRP-hydrogel groups was not statistically different from that inthe group treated with suture repair alone using the multiple groupcomparison model (ANOVA, Bonferroni Dunn post hoc testing p>0.04). Thedifference between the suture repair+PRP-hydrogel group and the intactligaments was also not significant (p>0.08). In contrast, the ligamentstreated with suture repair alone had a significantly lower energy tofailure than the intact ligaments (p<0.001).

The effect of platelet depletion on biomechanical parameters: Whenplatelet poor plasma was used in the collagen-platelet hydrogel, therewas no significant difference found between the collagen-PPP groups andthe suture repairs alone. This was true for all biomechanical parametersincluding the load at yield (p>0.50), maximum load (p>0.45),displacement at yield (p>0.70), stiffness (p>0.25) and energy to failure(p>0.39). The collagen-PPP group had significantly lower yield load,maximum load and stiffness when compared with the collagen-PRP group(p<0.006 for all comparisons).

TABLE 1 Biomechanical Properties of the ACL Four Weeks after Transectionand Repair Surgery/ Failure Energy Retr At B-L Load@Yield MaximumStiffness Displ@Yield to Fail Group completed jn (N) Load (N) (N/mm)(mm) (N * mm) Intact ACL n = 6 100%  142 +/− 38  179 +/− 37 48.6 +/−7.9  4.7 +/− 0.7 492 +/− 204 Suture Alone n = 7 0% 33 +/− 18  42 +/− 249.8 +/− 8.7 9.4 +/− 3.7 161 +/− 83  Suture + PPP n = 2 0% 19 +/− 4  25+/− 9 4.4 +/− 2.2 10.7 +/− 0.8  145 +/− 112 Suture + PRP n = 5 40%  93+/− 10 103 +/− 12 24.2 +/− 4.9  6.1 +/− 1.3 337 +/− 122 All valuesrepresent the mean +/− the standard deviation of the mean.

This study demonstrates that the biomechanical outcomes of strength andstiffness after primary repair of the ACL can be enhanced with use of acollagen-platelet rich hydrogel placed as a substitute provisionalscaffold in the ligament defect. This is a critical finding as priorresearch into stimulation of healing in articular tissue defects hasfocused on overcoming cellular deficiencies rather than scaffoldingdeficiencies. In this study, no cells (except the platelets and whiteblood cells contained in the platelet-rich plasma) were transplanted,yet a highly cellular repair tissue was seen within the defect afteronly four weeks. This suggests that at least in the ACL, there is asufficient intrinsic and/or extrinsic cellular response from theenvironment around the transected ACL to stimulate histologic healing ofthe defect if an appropriate scaffold is provided.

The advantages of this large animal model of complete ACL transectionand suture repair are multiple. The suture repair provides initialmechanical stability, and the use of absorbable suture that has minimalstrength at the end points of interest prevents the need for searchingthrough (and possibly destroying) the scar mass to release suture andallow for testing of the scar mass itself. Use of a large animal modelallows for easy identification of the structures of interest, both atthe time of ligament transection and retrieval, and ease of mechanicalrepair for surgeons versed in repair of human ligaments. Additionaltesting looking at the results of a complete ACL transection leftunrepaired for a period of time before surgical treatment would bebeneficial, as most patients will not be able to undergo immediaterepair; however, the costs of multiple animal surgeries and theadditional housing were beyond the finds available for this project.

The collagen-platelet rich hydrogel used here also has several majoradvantages over prior tissue engineered implants. There is no requiredcell or tissue harvest prior to implantation (other than phlebotomy).The collagen form used to mix with the platelet-rich plasma is similarto that used currently in plastic surgery procedures (Cooperman, L. S.;Mackinnon, V.; Bechler, G.; and Pharriss, B. B.: Injectable collagen: asix-year clinical investigation. Aesthetic Plastic Surgery, 9(2):145-51, 1985; Patel, M. P.; Talmor, M.; and Nolan, W. B.: Botox andcollagen for glabellar furrows: advantages of combination therapy.Annals of Plastic Surgery, 52(5): 442-7; discussion 447, 2004) where itis obtained either autologously or as a xenograft (Patel, M. P.; Talmor,M.; and Nolan, W. B.: Botox and collagen for glabellar furrows:advantages of combination therapy. Annals of Plastic Surgery, 52(5):442-7; discussion 447, 2004; Sclafani, A. P.; Romo, T., 3rd; Parker, A.;McCormick, S. A.; Cocker, R.; and Jacono, A.: Autologous collagendispersion (Autologen) as a dermal filler: clinical observations andhistologic findings. Archives of Facial Plastic Surgery, 2(1): 48-52,2000). Thus, future clinical application is likely to be relatively lowrisk, as opposed to treatment methods which require an additionalprocedure to procure cells for expansion (Adachi, N.; Sato, K.; Usas,A.; Fu, F. H.; Ochi, M.; Han, C. W.; Niyibizi, C.; and Huard, J.: Musclederived, cell based ex vivo gene therapy for treatment of full thicknessarticular cartilage defects. Journal of Rheumatology, 29(9): 1920-30,2002; Bellincampi, L. D.; Closkey, R. F.; Prasad, R.; Zawadsky, J. P.;and Dunn, M. G.: Viability of fibroblast-seeded ligament analogs afterautogenous implantation. J Orthop Res, 16(4): 414-20, 1998), or stemcells, or implanted recombinant growth factors or even viral vectors forgene therapy (Adachi, N.; Sato, K.; Usas, A.; Fu, F. H.; Ochi, M.; Han,C. W.; Niyibizi, C.; and Huard, J.: Muscle derived, cell based ex vivogene therapy for treatment of full thickness articular cartilagedefects. Journal of Rheumatology, 29(9): 1920-30, 2002; Evans, C. H.,and Robbins, P. D.: Genetically augmented tissue engineering of themusculoskeletal system. Clin Orthop, (367 Suppl): S410-8, 1999;Menetrey, J.; Kasemkijwattana, C.; Day, C. S.; Bosch, P.; Fu, F. H.;Moreland, M. S.; and Huard, J.: Direct-, fibroblast- andmyoblast-mediated gene transfer to the anterior cruciate ligament.Tissue Eng, 5(5): 435-42, 1999).

The use of Gelfoam as a carrier for the collagen-platelet hydrogel mayalso have contributed to the strength of the enhanced repairs. While wedid not run a control group with Gelfoam alone in this Example, thecollagen-PPP repairs were performed using Gelfoam and had mechanicalproperties inferior to that of the collagen-PRP group, suggesting thatthe platelets in the PRP group are more critical in stimulating healingof the ACL transection than the carrier itself. In addition, while thisis the first time healing of a complete transection of the ACL has beendemonstrated biomechanically, the recovery of biomechanical strength inthe defect remained incomplete at four weeks.

Example 3

In this example, we demonstrate biomechanical healing using a sponge,with anchor and suture in the absence of additional repair material/PRP.We conclude that biomechanical healing of the porcine ACL after completetransection and immediate suture repair using a collagen sponge is anovel treatment for this injury that is significantly better than thecurrent standard of care (ACL reconstruction).

Complete ACL transections were performed in five 30 kg Yorkshire pigsand repaired with a four stranded, absorbable suture repair using asuture anchor in the femur. In each animal, the repair was augmentedwith threading a collagen sponge onto the suture anchor before tying thesutures. No post-operative immobilization was used. The animals weresurvived for three months and then underwent in vivo magnetic resonanceimaging followed by euthanasia and immediate biomechanical testing. Sixcontrol knees with intact ACLs from three additional animals were usedas an intact ACL control group. The supplementation of suture anchorrepair with a collagen sponge resulted in formation of a large scar massin the region of the ACL. Load at yield, maximum load and ACL tangentmodulus were all significantly higher in the suture anchor repairsaugmented with collagen sponge than in ACL transections treated with thecurrent standard of care (ACL reconstruction) at the same time point.

Experimental Design

Five 30 kg female skeletally immature 4-month-old Yorkshire pigsunderwent ACL transection and suture anchor repair. All animals weretreated on one side with suture anchor repair augmented with collagensponge (n=5). All animals were euthanized after fourteen weeks. Justprior to euthanasia, the animals had in vivo MRI of both knees withgadolinium contrast to assess perfusion of the ACL wound site.Immediately after euthanasia, the knees were harvested biomechanicaltesting of the ACL complex performed as previously described. IntactACLs (n=6) from a separate group of age-matched, gender-matched andweight-matched animals were used as a control group for thebiomechanical studies.

Surgical Procedure:

Institutional Animal Care and Use Committee approvals were obtained forthis study prior to any surgical procedures. The pigs were pre-medicatedwith telazol 4.4-6.6 mg/kg IM, xylazine 1.1-2.2 mg/kg IM, and atropine0.04 mg/kg. They were intubated and placed on isoflurane 1-3% foranesthesia maintenance. After anesthesia had been obtained, the pigswere weighed and placed in the supine position on the operating roomtable. Both hind limbs were shaved, prepared with chlorhexidine followedby betadyne paint and sterilely draped. No tourniquet was used. Toexpose the ACL, a four-centimeter incision was made over the medialborder of the patellar tendon. The incision was carried down sharplythrough the synovium using electrocautery. The fat pad was released fromits proximal attachment and partially resected to expose theintermeniscal ligament. The intermeniscal ligament was released toexpose the tibial insertion of the ACL. A Lachman maneuver was performedprior to releasing the ACL to verify knee stability. Two #1 Vicrylsutures were secured in the distal ACL stump using a modified Kesslerstitch. The ACL was transected completely at the junction of the middleand proximal thirds using a No 12 blade. Complete transection wasverified visually and with a repeat Lachman maneuver that becamepositive in all knees with no significant endpoint detected aftercomplete transection. All knees were irrigated with sterile saline toremove synovial fluid before suture anchor placement. An absorbablesuture anchor (TwinFix AB 5.0 Suture Anchor with DuraBraid Suture(USP#2); Smith and Nephew, Inc, Andover Mass.) was placed at the back ofthe femoral notch. The knee was irrigated with 500 cc of sterile normalsaline to remove all synovial fluid. Hemostasis was carefully achievedusing pressure and a solution of 1:10,000 of epinephrine as needed. Oncehemostasis had been achieved, a collagen sponge was threaded ontosutures and up into the region of the proximal ACL stump in the notch.The sutures were tied with the knees in resting flexion (approximately70 degrees of flexion). The additional collagen sponge filled theintercondylar notch. The incisions were closed in multiple layers withabsorbable sutures.

The animals were not restrained post-operatively, and were allowed adlib activity. Once the animals recovered from anesthesia, they werepermitted to resume normal cage activity and nutrition ad lib. Buprenex0.01 mg/kg IM once and a Fentanyl patch 1-4 ug/kg transdermal wereprovided for post-operative analgesia. All animals were weight bearingon their hind limbs by 24 hours after surgery. After three months invivo, the animals were again anesthetized and underwent in vivo MRimaging using the protocol detailed below.

After the magnetic resonance images had been obtained, the animals wereeuthanized using Fatal Plus at 1 cc/10 lbs. No animals had any surgicalcomplications of difficulty walking normally, redness, warmth andswelling of the knee, fever or other signs of infection that would havenecessitated early euthanasia. The knees were retrieved and taken forimmediate ex vivo MR imaging and same-day biomechanical testing. Theknees were kept at 4 degrees C. until biomechanical testing and keptmoist using a saline spray and moist wraps.

The six intact control knees were obtained from age- gender- andweight-matched animals after euthanasia following surgical procedures tothe chest. The hind limbs were frozen at −20 degrees C. for three monthsand thawed overnight at 4 degrees C. before mechanical testing. Allother testing conditions for these knees were identical to those in theexperimental groups.

Magnetic Resonance Imaging: In vivo magnetic resonance imaging wasperformed at 1.5 Tesla (GE Medical Systems, Milwaukee, Wis.) with aneight-channel phased array coil at the specified time points. Scanningwas performed with the knees placed maximum extension (between 30 and 45degrees of flexion). Conventional MR included multiplane T1, FSE PD andT2 weighted images. Field of view (FOV): 16-18 cm, matrix: 256×256,(repetition time/echo time) TR/TE: 400/16, 2500/32, 3000/66 msec, echotrain length (ETL): 8, bandwidth (BW): 15 kHz, slice thickness: 3,interslice gap: 1 mm). Perfusion was evaluated by using spoiled gradientecho sequence (TR/TE=200/2 ms, flip angle=60, 3 mm slice thickness, and0.625 mm in plane resolution) with an intravenous contrast agent(Magnevist; Berlex, Wayne, N.J.) 0.2 ml/kg injected 10 s after the startof scan. Five images were obtained per slice, 78 s apart. Post contrastT1-weighted images were obtained (FOV: 16 cm, matrix: 256×256, TR/TE:400/9 msec, slice thickness: 3 mm, interslice gap: 1 mm) in the coronaland sagittal planes.

Biomechanical Testing: The bone-ligament-bone ACL complex from bothknees for each pig was tested in uniaxial tension as previouslydescribed. In brief, testing was performed with the knee flexed at 30degrees of flexion and at room temperature. Immediately afterpreconditioning, each specimen was tested to failure in uniaxial tensionat 20 mm/min. Close-range digital images were acquired at 3 Hz using ahigh resolution digital camera with a macro lens (PixeLINK PLA662Megapixel Firewire camera, PixeLINK, Ottawa ON, Canada) to determinefailure mode. The yield load, displacement at yield, tangent modulus(maximum slope of force-displacement curve), maximum load at failure,displacement at failure and total work to failure (area underforce-displacement curve) were determined from the force-displacementcurve measured for each bone-ligament-bone ACL complex. The yield loadrepresented the point along the normalized force-displacement curvewhere the mechanical behavior of the ACL complex departed from “linear”behavior and for the purposes of this analysis was defined as the pointwhere the tangent modulus declines by at least 2% from its maximumvalue. The displacement at yield was the displacement recorded at thissame point. The maximum load is the maximal normalized load sustained bythe ACL complex prior to failure and the displacement at failure thedisplacement recorded at the maximum load. The energy to failure wasderived by integrating the total area under the force-displacementcurve.

Results

MAGNETIC RESONANCE IMAGING: In vivo MRI demonstrated a progressivematuration of the repaired ACL from the large, bulky scar mass seen at 4weeks (Example 2) to an aligned structure with signal qualitiesindistinguishable from the normal ACL. The site of previous transectionof the ACL was no longer visible. The healing ACLs appeared moreorganized into tighter fascicles at the three month time point (FIG. 8).A synovial layer had been seen to form over the ligaments, and bloodvessels were seen on the surface of the ligaments. FIG. 8: shows thegross appearance of the Intact ACL (8A) and repaired ACL (8B) at threemonths (arrows). Of note is the fascicular organization of the tissue onthe left.

Mechanical Properties: The strength of the repairs using suture andcollagen sponge averaged 52% of the intact ACL strength at the threemonth time point. This is favorable in comparison with the strength ofACL reconstruction in animal models, where the strength at three and sixmonths is only approximately 20% of the intact ACL (FIG. 9).

The stiffness of the suture anchor/collagen sponge repairs was 36% thatof the intact ACL—this also compares favorably with the current standardof care (ACL Reconstruction) where the stiffness at 12 weeks is only 23%of the intact ACL (FIG. 9).

FIG. 9 shows biomechanical properties of Suture Anchor/Sponge Repair vsthe current standard of care for ACL injuries (ACL Reconstruction orACLR, ACLR data from Hunt et al, 2005) at 3 months in vivo. All valuesare normalized by the properties of the intact ACL in the specificanimal model to compensate for variation in animal size and anatomy. Thestrength of the primary repaired ligaments is more than three times ashigh as the ACL Reconstructed knees.

This Example demonstrates that the biomechanical outcomes of strengthafter primary repair of the ACL transection can be enhanced with thenovel technique of a collagen sponge threaded on the suture anchorsutures and thus located within the repair site. The strength at threemonths after repair is over 50% of the normal ACL strength—a value morethan twice as high as the strength of ACL reconstruction at similar timepoints. In summary, use of a collagen sponge can stimulate biomechanicalhealing after suture anchor repair. The data supports significantchanges in our clinical approach to ACL rupture, from resection andreplacement towards repair and regeneration. ACLR data from Hunt et al,2005

The foregoing written specification is considered to be sufficient toenable one skilled in the art to practice the invention. The presentinvention is not to be limited in scope by examples provided, since theexamples are intended as a single illustration of one aspect of theinvention and other functionally equivalent embodiments are within thescope of the invention. Various modifications of the invention inaddition to those shown and described herein will become apparent tothose skilled in the art from the foregoing description and fall withinthe scope of the appended claims. The advantages and objects of theinvention are not necessarily encompassed by each embodiment of theinvention. Those skilled in the art will recognize, or be able toascertain using no more than routine experimentation, many equivalentsto the specific embodiments of the invention described herein. Suchequivalents are intended to be encompassed by the following claims.

All references disclosed herein are incorporated by reference in theirentirety.

1.-86. (canceled)
 87. A construct for repair or augmentation of a tissueexposed to synovial fluid comprising a compressible porous supportscaffold configured for repair or augmentation of the tissue, whereinthe scaffold allows cell ingrowth.
 88. The construct of claim 87,wherein the tissue is a ligament, tendon, meniscus, cartilage, labrum ormuscle.
 89. The construct of claim 88, wherein the ligament is an ACL.90. The construct of claim 87, wherein the scaffold is expandable. 91.The construct of claim 87, further comprising a suture.
 92. Theconstruct of claim 91, wherein the suture is bioabsorbable.
 93. Theconstruct of claim 87, further comprising an anchor.
 94. The constructof claim 93, wherein the anchor is selected from the group consisting ofa screw, a barb, a helical anchor, a staple, a clip, a snap, and arivet.
 95. The construct of claim 87, further comprising a boneinsertion device.
 96. The construct of claim 87, wherein the scaffoldcomprises a therapeutic agent.
 97. The construct of claim 5, wherein thetherapeutic agent is selected from the group consisting of hormones,cytokines, growth factors, clotting factors, anti-protease proteins,angiogenic proteins, antiangiogenic proteins, bone morphogenic proteins,osteoinductive factor, fibronectin, endothelial cell growth factor,cementum attachment extracts, ketanserin, human growth hormone, animalgrowth hormones, epidermal growth factor, interleukin-1, human alphathrombin, transforming growth factor, insulin-like growth factor,platelet derived growth factors, fibroblast growth factors, andperiodontal ligament chemotactic factors.
 98. The construct of claim 87,wherein the scaffold comprises a clotting factor.
 99. The construct ofclaim 87, wherein the scaffold comprises platelets or platelet richplasma.
 100. The construct of claim 87, wherein the scaffold comprisescollagen.
 101. The construct of claim 87, wherein the scaffold is asponge.
 102. The construct of claim 87, wherein the scaffold is tubular,semi-tubular, or cylindrical.
 103. The construct of claim 87, whereinthe scaffold is an amorphous shape that conforms to the shape of arepair space.
 104. The construct of claim 87, comprising the scaffold,at least one bone insertion device and at least one suture.
 105. Theconstruct of claim 104, comprising a platelet and a clotting factor inthe scaffold.
 106. The construct of claim 87, wherein the constructfurther comprises xenograft tissue, allograft tissue, or autografttissue.
 107. A kit comprising the construct of claim 87, and a scaffold,suture, and a bone insertion device.
 108. A kit comprising the constructof claim 87, a scaffold and platelets or means for obtaining plateletsfrom a patient.
 109. The kit of claim 107 or 108, further comprising atherapeutic agent.
 110. The kit of claim 109, wherein the therapeuticagent is a clotting factor.